The long-term objectives are to understand the cellular mechanisms of transduction and frequency selectivity in auditory hair cells. The work will focus on the roles of intracellular Ca2+, especially those pertinent to regulation of the electrical resonance mechanism that tunes the receptor potentials of hair cells in many lower vertebrates. The electrical resonance stems from the interplay of a voltage-dependent Ca2+ current and a Ca2+-activated K+ current. However, the cellular mechanism which dictates a hair cell's tuning characteristics according to the cell's location on the basilar membrane is not known. The experiments will examine the attributes of single hair cells of the turtle's and alligator's cochleas employing patch-clamp recording and imaging of signals from Ca2+- sensitive dyes. The specific aims during this period are: (1) to map the ionic currents in hair cells along the cochlea, testing whether cells tuned to higher frequencies have more voltage-dependent Ca2+ channels and faster activating K+ channels; (2) to characterize the size and kinetics of single K+ channels in membrane patches and explore their modulation by factors such as internal Ca2+ and protein phosphorylation; (3) using Ca2+-sensitive dyes, to measure the metabolism of intracellular Ca2+, its resting concentration and its entry and extrusion mechanisms. Does the resting concentration vary with the frequency of tuning of the hair cell? (4) by manipulating its intracellular concentration, to examine calcium's involvement in adaptation of the mechano-electrical transduction channels and its link to active motion of the hair bundle; differences in mechanical properties will be sought between tall and short hair cells of the alligator's cochlea. All such experiments will involve measuring the mechanics of the hair bundles by displacing them with a flexible fibre of known compliance. Information from these studies may lead to insights into the mechanisms controlling the expression and localization of membrane channels which may be a target for malfunction in disease. It may also reveal a stringent demand on the intracellular Ca2+ level for channel regulation and functioning of the hair cells. The mechanisms mediated by internal Ca2+ are probably common to all hair cells, including those in the mammalian cochlea, and they may be the sites at which irreversible damage occurs during acoustic over-stimulation or poisoning with aminoglycoside antibiotics.